Marfan syndrome is characterized by high expression of matrix metalloproteinases (MMPs) in aortic smooth muscle cells (AoSMCs) associated with medial elastolysis and aortic root aneurysm. We aimed to ...reduce aortic elastolysis through decrease of MMP expression with decoy oligodeoxynucleotides (dODNs) neutralizing the transcription factor activating factor-1 (AP-1). AP-1 abundance in nuclear extracts as well as MMP-2 and MMP-9 expression were significantly increased in isolated mAoSMC of mgR/mgR Marfan mice compared to wild-type cells. Exposure to AP-1 neutralizing dODNs resulted in a significant reduction of basal and interleukin-1β-stimulated MMP expression and activity in mAoSMCs. Moreover, increased migration and formation of superoxide radical anions was substantially decreased in mAoSMCs by AP-1 dODN treatment. Aortic grafts from donor Marfan mice were treated with AP-1- dODN ex vivo and implanted as infrarenal aortic interposition grafts in mgR/mgR mice. Pretreatment of aortic grafts with AP-1 dODN led to reduced elastolysis, macrophage infiltration, and MMP activity. Permeability of the endothelial monolayer was increased for dODN in mgR/mgR aortae with observed loss of tight junction proteins ZO-1 and occludin, enabling dODN to reach the tunica media. Targeting AP-1 activity offers a new potential strategy to treat the vascular phenotype associated with Marfan syndrome.
Cardiac allograft vasculopathy (CAV) is the main obstacle for long-term survival after heart transplantation. Alloimmune mediated chronic vascular rejection results in several mechanisms like ...platelet activation, immigration of inflammatory cells through the endothelial layer and proliferation and migration of smooth muscle cells (SMCs). Serotonin (5-HT) promotes these processes via activation of 5-HT2 receptors. We hypothesized that inhibiting 5-HT2 receptors ameliorates the development of CAV.
CBA/JRj mice recieved aortic grafts from C57BL/6 mice. After transplantation until recovery of organs, recipients were treated with serotonin receptor antagonists: sarpogrelate (5-HT2A), SB 204741 (5-HT2B) or terguride (5-HT2A+B). Mice were sacrificed after 14 days for qRT-PCR analysis or after 30 days for histological evaluation. Serum serotonin ELISA was done at both time points.
Elevated serum serotonin levels were significantly reduced after 5-HT2A antagonist treatment as was 5-HT2A receptor expression. This went along with reduced inflammation characterized by significantly fewer infiltrating macrophages and pro-inflammatory intragraft cytokines and with reduced tissue remodeling evident as significantly less neointima formation.
Inhibition of the 5HT/5-HT2A receptor axis leads to significantly reduced neointima proliferation after aortic transplantation associated with reduced transendothelial migration of macrophages and decreased expression of inflammatory cytokines. These findings have translational implications as inhibitors of 5HT2A like sarpogrelate are already approved for clinical use.
•5HT2A receptor inhibitors reduce neointima proliferation after vessel transplantation•They may reduce migration of macrophages and expression of inflammatory cytokines•These findings have translational implications as sarpogrelate is clinically approved
Abstract Background Every transplanted organ relies on a reliable and sound vascular system. Therefore, our study focused on the investigation if platelet inhibition alone or combined with ...mTOR-inhibition has a beneficial effect on the microvascular integrity in allogeneic murine skin grafts. Methods Skin transplantation was performed from fully MHC-mismatched C57BL/6 (H-2b ) donors to CBA/J (H-2k ) recipient mice. Skin allograft recipients were assigned to several experimental groups and either treated with clopidogrel alone, everolimus alone or a combination of both. Graft survival was evaluated and transplants were harvested after 8 days and analyzed for CD31 and C4d by immunohistochemistry. Results Untreated allografts showed a reduced amount of CD31 on postoperative day 8 as well as an increase in C4d compared to isografts. All treated animals showed a significant improvement regarding CD31 1577.7 ± 200.4 (clopidogrel)/1702.8 ± 151.1 (clopidogrel + everolimus) vs. 479.7 ± 184.2 (control), n = 8, p < 0.05 and C4d 420.9 ± 70.9 (clopidogrel)/324.5 ± 77.3 (clopidogrel + everolimus) vs. 772.4 ± 159.7 (control), n = 8, p < 0.05. In addition, skin grafts of animals treated with clopidogrel and everolimus survived significantly longer compared to untreated controls 19.2 ± 4.2 d vs. 12.8 ± 2.4 d, n = 10, p < 0.05. Conclusion In this study we could show that clopidogrel alone and in combination with everolimus substantially improved microvascular integrity and resulted in increased survival time of skin grafts.
Nintedanib is a small molecule tyrosine kinase inhibitor that blocks the action of the platelet-derived growth factor receptor (PDGFR), the vascular endothelial growth factor receptor (VEGFR) and the ...fibroblast growth factor receptor. All of these receptors have been shown to be involved in the development of cardiac allograft vasculopathy (CAV) after heart transplantation. We therefore hypothesized that blocking these tyrosine kinase receptors with nintedanib could prevent CAV.
CBA/JRj (H2
) mice underwent an abdominal aortic transplantation with a graft derived from fully allogeneic C57BL/6JRj (H2
) mice. Nintedanib was given daily from the first day after transplantation until harvest on day 14 for polymerase chain reaction analysis of intragraft cytokine expression or harvest on day 30 for histological analysis of the graft.
Nintedanib treatment resulted in significantly reduced neointima formation in the aortic graft compared with untreated control allografts. Interestingly, the immigration of smooth muscle cells into the neointima was markedly reduced while graft infiltrating macrophages and T cells were not altered in nintedanib-treated animals. The expression of the growth factor PDGF was significantly reduced in the nintedanib group going along with a distinctly reduced expression of the corresponding receptors PDGFR α and -β.
Treatment with nintedanib caused a significant reduction of CAV development after aortic transplantation in mice. We hypothesize the attenuated neointima formation in nintedanib-treated animals to be mediated by a direct inhibition of intimal smooth muscle cell proliferation via reduced expression of PDGF and the appropriate receptors PDGFR α + β.
Abstract Background Obliterative bronchiolitis (OB) is the major limiting factor for long-term survival after lung transplantation. As previously shown, donor treatment with a PHD-inhibitor ...activating hypoxia-inducible transcription factors (HIFs) prevents graft injury both in an allogenic kidney and aortic allograft transplant model. The aim of this study was to investigate the effect of HIF activation with a PHD-inhibitor on the development of OB. Methods Fully MHC-mismatched C57BL/6 (H-2b ) donor tracheas were orthotopically transplanted into CBA/J (H-2k ) recipients. Donor animals received a single dose of PHD-inhibitor 2-(1-chloro-4-hydroxyisoquinoline-3-carboxamido) acetate (ICA) (40mg/kg i.p.) or vehicle control 4h before transplantation. Animals were harvested 14, respectively 30 days after transplantation and grafts were analyzed by histology, mRNA expression and immunofluorescence. Results Donor pre-conditioning with ICA resulted in HIF accumulation and induction of HIF target genes: HO-1, VEGF, MIF, TGFβ, and EpoR, which persisted during different times of ischemia. Vehicle treated controls showed substantial luminal obliteration on postoperative day 30 in contrast to groups pre-treated with ICA luminal obliteration 29.2 + 5% (ICA) vs. 36.7 + 8% (control), p < .01. We found significantly lower expression of TNFα, PDGFß, MCP-1, E-selectin, and ICAM-1 14 days after ICA premedication. In addition ICA pre-treated groups revealed decreased T-cell and macrophage infiltration in tracheal grafts on days 30 after transplantation (p < .05). Conclusion Pre-treatment with ICA effectively reduced obliterative bronchiolitis. Our data suggest that activation of hypoxia-inducible transcription factors (HIFs) and thereby adaptation to low oxygen prevents the development of OB and allograft injury. Pharmaceutical inhibition of PHDs appears to be an attractive strategy for organ preservation that deserves further clinical evaluation.
Monotherapy with clopidogrel reduced the formation of transplant arteriosclerosis in a murine aortic allograft model. However, the underlying immunologic mechanisms are still unknown.
Fully major ...histocompatibility complex-mismatched C57BL/6 (H2b) donor aortas were transplanted into CBA.J (H2k) recipients and mice received different doses (1, 10, and 20 mg/kg) of clopidogrel, an antagonist of the P2Y12 ADP receptor on platelets, or control saline for 30 days. Blood was analyzed for changes in adhesion molecule and sCD40L concentrations by ELISA. Grafts were analyzed by histology, morphometry, and immunofluorescence on day 30 after transplantation. Intragraft cytokine mRNA production was analyzed by reverse-transcriptase polymerase chain reaction on day 14 after transplantation.
Treatment with clopidogrel resulted in significantly decreased blood concentrations of sCD40L and P-selectin after transplantation. Cellular analysis of the aortic transplant revealed fewer numbers of infiltrating dendritic cells (CD205+) and macrophages (F4/80+) after application of clopidogrel, whereas T-cells within the graft were unaltered. In addition cellular P-/E-selectin, ICAM-1, and platelet-derived-growth-factor (PDGF)-beta surface expression were significantly reduced as compared with untreated controls. Intragraft mRNA expression confirmed these results and showed significant lower production of P-/E-selectin, ICAM-1, and PDGF-beta after treatment with clopidogrel. Antiglycoprotein-Ib and antiglycoprotein VI had no beneficial effect on the development of transplant arteriosclerosis.
This report shows that application of clopidogrel after transplantation results in a reduction in adhesion molecule expression within the blood and transplant tissue and is associated with reduced transendothelial migration of dendritic cells and macrophages within the vascular wall.
Abstract The mTOR inhibitor everolimus (EVL) can be used for calcineurin inhibitor-sparing immunosuppression in heart transplantation (HTx). However, comparable data regarding clinical outcomes in ...HTx recipients receiving EVL either with dosage reduction of cyclosporine A (CSA) or with dosage reduction of tacrolimus (TAC) is scarce. In a retrospective data analysis, we compared 5-year clinical outcomes in 154 maintenance patients receiving EVL with CSA (n = 106) or TAC (n = 48). The primary endpoint was a composite of death, graft loss and EVL discontinuation (treatment failure). Secondary endpoints were kidney function, cardiac rejection, cytomegalovirus infection and biochemical safety parameters. In the CSA and TAC group, the primary endpoint was reached by 59.8% and 53.1%, respectively (P = 0.716). Five-year mortality was 30.4% (CSA group) and 23.13% (TAC group), respectively (P = 0.371), and freedom from EVL discontinuation was 53.3% and 59.6% (P = 0.566) in the respective groups. Covariate-adjusted relative risk of treatment failure was in the CSA group = 1.28 (95% CI: 0.70–2.34; P = 0.43) compared with the TAC group. The course of covariate-adjusted estimated glomerular filtration rate and freedom from cytomegalovirus infection was similar in the two groups (P = 0.502 and P = 0.476), whereas covariate-adjusted freedom from rejection was lower in the CSA group compared with the TAC group (P = 0.023). Lipid status and blood cell counts were comparable between groups. In conclusion, data indicate that EVL plus reduced TAC is not superior to EVL plus reduced CSA regarding treatment failure and kidney function. However, compared with EVL plus reduced CSA, EVL plus reduced TAC seems to reduce cardiac rejections.
OBJECTIVES
The development of transplant arteriosclerosis, the hallmark feature of heart transplant rejection, is associated with a chronic immune response and also influenced by an initial injury to ...the graft through ischaemia and reperfusion. Hypoxia-inducible transcription factor (HIF)-1 pathway signalling has a protective effect against ischaemia–reperfusion injury and has already been demonstrated to ameliorate allograft nephropathy in previous animal studies. Therefore, the aim of this study was to investigate the effect of stabilization of hypoxia-inducible transcription factors with a prolyl-hydroxylase domain (PHD) inhibitor on transplant arteriosclerosis in an experimental aortic allograft model.
METHODS
MHC-class I mismatched C.B10-H2(b)/LilMcdJ donor thoracic aortas were heterotopically transplanted into the abdominal aorta of BALB/c mice. Donor animals received a single dose of the PHD inhibitor 2-(1-chloro-4-hydroxyisoquinoline-3-carboxamido) acetate (ICA) (40 mg/kg) or vehicle i.p. 4 h before transplantation. Intragraft HIF accumulation after ICA treatment was detected by immunohistochemistry before and after cold ischaemia (n = 5). Grafts were harvested 30 days after transplantation and analysed by histology (n = 7) and immunofluorescence (n = 7). In addition, intragraft mRNA expression for cytokines, adhesion molecules and growth factors was determined on Day 14 (n = 7).
RESULTS
Donor preconditioning with ICA resulted in HIF accumulation in the aorta and induction of the HIF target genes vascular endothelial growth factor and transforming growth factor-beta. Vascular lesions were present in both experimental groups. However, there was significantly reduced intimal proliferation in preconditioned grafts when compared with vehicle controls intimal proliferation 31.3 ± 8% (ICA) vs 55.3 ± 20% (control), P < 0.01. In addition, experimental groups revealed a down-regulation of E-selectin (−57%) and MCP1 (−33%) expression after ICA pretreatment compared with controls, going along with decreased T-cell 1.4% CD4+ T-cell infiltration vs 8.4% (control) and 4.9% CD8+ T-cell infiltration vs 10.7% (control), dendritic cell (0.6% dendritic cells infiltration vs 1.9% infiltration(control) and macrophage infiltration 4.8% macrophages (ICA) vs 10.9% (control) within vascular grafts.
CONCLUSIONS
These data of an animal transplant model show that the pharmaceutical activation of HIF with endogenous up-regulation of protective target genes leads to adaptation of the graft to low oxygen-saturation and hereby attenuates the development of transplant arteriosclerosis and allograft injury. Pharmaceutical inhibition of PHDs appears to be a very attractive strategy for organ preservation that deserves further clinical evaluation.
Heart transplant recipients are at increased risk of developing malignant neoplasms. Administration of the calcineurin inhibitors cyclosporine A (CSA) or tacrolimus (TAC) may contribute to this risk.
...We compared tumor incidence in heart transplant recipients receiving either CSA (n=25) or TAC (n=120) as maintenance immunosuppressive therapy. Exclusion criteria were therapy with mammalian target of rapamycin-inhibitors, death within the first postoperative year, re-transplantation, and age less than 18 years.
The 2 study groups were comparable with respect to sex, primary and concomitant diagnoses, and mean follow-up (60.7 ± 19.3 months in the CSA group vs. 59.8 ± 18.1 months in the TAC group; P=0.81). The CSA group was, however, significantly older compared with the TAC group (58.8 ± 11.4 years vs. 49.1 ± 13.0 years, P=0.001), as was the donor age of the CSA group (43.2 ± 11.2 years vs. 37.0 ± 11.7 years, P=0.02). In the CSA group, 5 patients (20%) developed malignant neoplasms compared with 10 patients (8.3%) in the TAC group (P=0.14). Covariate-adjusted 5-year tumor-free survival was comparable between groups (relative risk for the CSA group =1.162 95% CI: 0.378-3.572; P=0.794). Moreover, covariate-adjusted 5-year overall survival did not differ between the 2 groups (relative risk for the CSA group =1.95 95% CI: 0.53-7.19; P=0.36). The incidence of infection, acute rejection, graft vasculopathy, renal failure, and neurological complications was also comparable between the 2 groups.
Our data indicate that tumor incidence does not significantly differ in patients receiving CSA or TAC as maintenance therapy.
Clopidogrel reduces post‐transplant obliterative bronchiolitis Preidl, Raimund H. M.; Eckl, Sebastian; Ramsperger‐Gleixner, Martina ...
Transplant international,
October 2013, 2013-Oct, 2013-10-00, 20131001, Letnik:
26, Številka:
10
Journal Article
Recenzirano
Odprti dostop
Summary
Survival after lung transplantation is mainly limited by the development of chronic lung allograft dysfunction (CLAD). The aim of this study was to investigate if platelet inhibition by ...clopidogrel has an influence on the formation of obliterative bronchiolitis, the histopathological correlate to bronchiolitis obliterans syndrome, present in the majority of patients suffering from CLAD. C57Bl/6(H2b) donor tracheas were orthotopically transplanted into CBA.J(H2k). Mice received different doses of clopidogrel alone or in combination with tacrolimus or everolimus. Grafts were analyzed by histology and immunofluorescence method on postoperative days 15, 30 or 60. Cytokines were analyzed by real‐time polymerase chain reaction on postoperative day 21 and alloantibodies by FACS. Mice treated with 20 mg/kg/day clopidogrel for 30 days showed reduced obliteration 34.40 ± 3.76% (20 mg/kg/day clopidogrel) vs. 49.92 ± 2.11% (control), n = 5, P < 0.05. Platelet inhibition resulted in significant lower infiltration of T cells and macrophages, and we also found significantly lower expression of IL‐12, IL‐4, IL‐6, TNF‐α, TGF‐β, PDGFβ, MCP1, P‐/E‐selectin, ICAM1 and CD40L after treatment with clopidogrel. Combination of 1 mg/kg/day clopidogrel and 0.05 mg/kg/day everolimus or 12 mg/kg/day tacrolimus revealed a synergistic effect. Humoral immunity as manifested by donor‐specific alloantibody secretion was also impaired after treatment with clopidogrel. Here, we can show that platelet inhibition by clopidogrel as a single treatment and in combination with tacrolimus or everolimus reduced the development of fibrosis and obliteration in tracheal allografts.
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